Angiogenin-Induced RNA Cleavage in Cancer

描述（由申请人提供）：本提案的目的是确定血管生成素（ANG）诱导的转移RNA （tRNA）切割如何促进肿瘤生长和存活。我们的主要假设是，ang诱导的tRNA切割改变了蛋白质翻译，从而增强了促血管生成、促生长和促生存蛋白的表达，从而促进了肿瘤的发生。这是基于我们自己的初步数据，表明ANG选择性地切割trna以产生抑制翻译起始的生物活性片段（即tirna）。这项研究的基本原理是，一旦我们知道tRNA切割如何重新编程蛋白质翻译，我们将能够调节这一事件来治疗癌症。我们将通过完成三个具体目标来检验我们的中心假设：确定YB-1如何与核糖核酸协同抑制癌细胞的翻译起始。我们的工作假设是，核糖核酸结合到YB-1的核酸结合冷冲击结构域，促进与翻译起始复合物组分的相互作用。目标2。确定ang诱导的tRNA切割如何改变蛋白质翻译以增强癌细胞的增殖和存活。我们的工作假设是，tiRNAs“激活”YB-1，允许优先翻译内部核糖体进入位点（IRSE）和上游开放阅读框(uORF)-含有编码促进肿瘤细胞生长和存活的蛋白质的转录本。目标3。确定ang诱导的tRNA切割在肿瘤发生中的作用。我们的工作假设是，tiRNAs在ANG的下游作用，促进肿瘤生长。我们将确定YB-1，一个靶向翻译机制以促进上皮-间质转化的癌基因，是否与tRNA片段合作，重新编程癌细胞中的蛋白质翻译。我们将使用无偏见的基因阵列和候选基因方法来鉴定转录本，这些转录本的翻译受到tRNA切割的调节。我们将使用siRNA敲低来确定ANG抑制剂RNH1是否具有肿瘤抑制蛋白的功能。我们将确定tRNA片段、YB-1和RNH1的表达如何影响小鼠异种移植物的肿瘤发生。最后，我们将确定tRNA切割是否作为前列腺癌的预后生物标志物。提出的研究的贡献将是确定ANG的核糖核酸酶活性如何促进肿瘤细胞的生长和存活。这一贡献是重要的，因为它为开发阻止ang介导的肿瘤生长的药理学策略提供了分子基础。这项研究具有创新性，因为它专注于ANG的直接靶点，即其RNA底物，并试图确定tRNA切割如何促进肿瘤发生。

DESCRIPTION (provided by applicant): The objective of this proposal is to determine how angiogenin (ANG)-induced transfer RNA (tRNA) cleavage facilitates tumor growth and survival. Our central hypothesis is that ANG-induced tRNA cleavage alters protein translation to enhance the expression of angiogenic, pro-growth and pro-survival proteins to promote tumorigenesis. This is based upon our own preliminary data showing that ANG selectively cleaves tRNAs to produce bioactive fragments (i.e., tiRNAs) that inhibit translation initiation. The rationale for te proposed research is that, once we know how tRNA cleavage re-programs protein translation, we will be able to modulate this event to treat cancer. We will test our central hypothesis by the completion of three specific aims: AIM 1. Determine how YB-1 cooperates with tiRNAs to inhibit translation initiation in cancer cells. Our working hypothesis is that tiRNAs bind to the nucleic acid-binding cold shock domain of YB-1 to promote interactions with components of the translation initiation complex. AIM 2. Determine how ANG-induced tRNA cleavage alters protein translation to augment the proliferation and survival of cancer cells. Our working hypothesis is that tiRNAs "activate" YB-1 to allow the preferential translation of internal ribosome entry site (IRSE) and upstream open reading frame (uORF)-containing transcripts encoding proteins that promote tumor cell growth and survival. AIM 3. Determine the role of ANG-induced tRNA cleavage in tumorigenesis. Our working hypothesis is that tiRNAs act downstream of ANG to promote tumor growth. We will determine whether YB-1, an oncogene that targets the translational machinery to promote the epithelial- mesenchymal transition, partners with tRNA fragments to re-program protein translation in cancer cells. We will use non-biased gene array and candidate gene approaches to identify transcripts whose translation is modulated in response to tRNA cleavage. We will use siRNA knockdown to determine whether the ANG inhibitor RNH1 functions as a tumor suppressor protein. We will determine how the expression of tRNA fragments, YB-1 and RNH1 influence tumorigenesis in murine xenografts. Finally, we will determine whether tRNA cleavage serves as a prognostic biomarker for prostate cancer. The contribution of the proposed research will be to determine how the ribonuclease activity of ANG promotes the growth and survival of tumor cells. This contribution is significant because it provides a molecular basis for the development of pharmacologic strategies to prevent ANG-mediated tumor growth. The proposed research is innovative because it focuses on the direct target of ANG, its RNA substrates, and attempts to determine how tRNA cleavage promotes tumorigenesis.